A novel material for the improvement of HIT detection

ABSTRACT

Background: Heparin-induced thrombocytopenia (HIT) is a drug reverse disorder (happening in up to 5% of patients) that could lead to lethal consequences. The patient is confirmed with HIT if HIT antibodies (Abs) are present. Recently, HIT Abs were detected also in clinically ill Covid-19 and VITT patients. Early detection of HIT is crucial, but the widely used immunoassays provide only ~50% accuracy while functional assays required fresh platelets, which can only be performed in limited laboratories. Thus, the development of a simple method but accurate detection for confirmation of HIT Abs is highly desired. Aim(s) We aim to discover cell membranes as novel materials to improve the detection of HIT Abs. Method(s) We used the laser scanning confocal microscope (LSCM) to visualize and confirm the binding of HIT Abs (including monoclonal Abs and patient's sera) on breast cancer cell line MDA-MB- 231. Flow cytometry was used to quantify the binding of antibodies via bound PF4 or PF4/Heparin complexes on the cells. Optical density caused by bindings of HIT Abs to cells was detected by integrating the cells into the ELISA setup. Result(s) We found that the MDA-MB- 231 cell line allows bindings of HIT Abs via platelet factor 4 (PF4) or PF4/Heparin complexes. Bound PF4 on cells allowed better binding of HIT Abs than the bound PF4/H complexes. Heparin inhibits HIT antibody bindings to cells. Importantly, non-HIT Abs also bind to the cells through PF4 but are much weaker than the binding of the HIT Abs. The different binding patterns between HIT and non-HIT Abs could be clearly distinguished. Conclusion(s) The MDA-MB- 231 cell line has a great potential to distinguish HIT from non-HIT Abs through PF4 but not PF4/H complexes. (Figure Presented).